The generation of monoclonal antibodies against human peroxisome proliferator-activated receptors (PPARs).

Toshiya Tanaka, Tetsu Takeno, Yuichiro Watanabe, Yasutoshi Uchiyama, Takeshi Murakami, Hisahiko Yamashita, Akifumi Suzuki, Rie Aoi, Hiroko Iwanari, Shu Ying Jiang, Makoto Naito, Keisuke Tachibana, Takefumi Doi, Andrew I. Shulman, David J. Mangelsdorf, Raphael Reiter, Johan Auwerx, Takao Hamakubo, Tatsuhiko Kodama

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

Monoclonal antibodies (Mabs) are valuable reagents for the purification, characterization and immunolocalization of proteins.In this study, we raised Mabs against human peroxisome proliferator-activated receptors (PPARs) using baculovirus particles displaying surface glycoprotein gp64-fusion proteins as the immunizing agent.In this system, to display fusion proteins on the viral surface, the amino terminal sequences of human PPARd and PPARg2 are inserted in-frame between the signal sequence and the mature domain of the gp64 nucleotide sequence.Mabs were raised by immunization with whole virus without a purification of the target antigens.The Mabs generated by this novel method were shown to recognize not only the gp64-PPARs fusion protein, but also mature, expressed proteins by a wide variety of techniques, including immunohistochemistry, immunoblotting, and electrophoretic mobility shift assays (EMSAs).Transfection of the transfer vector containing a nucleotide sequence encoding less than 30 amino acids along with linearized baculovirus DNA allows for the production of a high affinity antibody against the corresponding mature form.This method is of potential utility in that it allows the production of valuable antibodies without the requirement of a protein purification step.

Original languageEnglish (US)
Pages (from-to)233-242
Number of pages10
JournalJournal of atherosclerosis and thrombosis
Volume9
Issue number5
DOIs
StatePublished - 2002

ASJC Scopus subject areas

  • Internal Medicine
  • Cardiology and Cardiovascular Medicine
  • Biochemistry, medical

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    Tanaka, T., Takeno, T., Watanabe, Y., Uchiyama, Y., Murakami, T., Yamashita, H., Suzuki, A., Aoi, R., Iwanari, H., Jiang, S. Y., Naito, M., Tachibana, K., Doi, T., Shulman, A. I., Mangelsdorf, D. J., Reiter, R., Auwerx, J., Hamakubo, T., & Kodama, T. (2002). The generation of monoclonal antibodies against human peroxisome proliferator-activated receptors (PPARs). Journal of atherosclerosis and thrombosis, 9(5), 233-242. https://doi.org/10.5551/jat.9.233